The long-term goal of this research is to develop and apply molecular genetic techniques for the control of arthropod-borne virus (arbovirus) diseases. Conventional control measures, including pesticide application, environmental sanitation, and immunization have proven to be effective in short-term reduction of arbovirus diseases, but not in long-term reduction or eradication. Indeed, many arbovirus disease remain significant pathogens or are resurgent in previously controlled areas. In this proposal, novel molecular genetic strategies will be investigated for manipulation of the Aedes triseriatus genome to intracellularly immunize the mosquito to prevent to reduce LaCrosse (LAC) virus infection and transmission. The specific aims are: 1. Intracellular immunization: The efficacy of developed antiviral constructs, which express specific virus proteins, virus antisense sequences, virus specific ribozymes, will determined using a Sindbis virus expression system. The Sindbis system permits expression of antiviral constructs in vitro in cell cultures and in vivo in mosquitoes. Constructs that interfere with virus infection, replication, and/or transmission will then be candidates for generation of transgenic mosquitoes. 2. The Aedes densonucleosis mosquitoes. Expression of transform mosquitoes by either genomic integration or as a stable episome will be tested. 4. An arbovirus refractory mosquito will be bioengineered by introducing promising antiviral constructs into the genome by transduction or other transformation procedures. In addition to providing important information about the potential of intracellular immunization for control of arbovirus (and other virus) disease, these studies will provide invaluable information about gene regulation and expression in vectors, and the potential for utilization of the new Aedes parvovirus as a tool for genetic manipulation of vectors. Considerable information should be forthcoming from these proposed studies which could provide novel strategies for control of these very important vecto-borne virus pathogens of humans and animals.